Selection of probiotics and prebiotics for synbiotics and confirmation of their in vivo effectiveness

Abstract

Influence of fructan-type oligosaccharides (as prebiotics) on growth and acidifying activity of Bifidobacterium strains (as probiotics) was studied in vitro, using minimal nutrition media. The selected synbiotic pairs of stimulated bifidobacteria strains and oligosaccharides enhancing their growth were studied in vivo to determine the effect of probiotics, prebiotics and synbiotics on microecology of gut. Daily, >10⁹ live cells of bifidobacteria strains and/or 5% (w/w) of oligofructose in the diet were orally administered to Wistar rats. After a 14-day feeding experiment, the numbers of faecal bifidobacteria, aerobic and facultatively anaerobic bacteria, coliforms, and spores of anaerobic and aerobic bacteria were determined using microbiological methods. In vitro studies showed that the majority of Bifidobacterium species utilised FOS and low-polymerised inulins, but only 18 out of 30 strains tested (mostly of B. longum and B. animalis species) were stimulated. Incorporation of oligofructose into the diet stimulated the proliferation of faecal bifidobacteria by 1.6 log cfu/g in comparison to the control. B. longum KN29.1 and KNA1 selected in vitro, slightly increased the faecal bifidobacteria live cell number by about 0.6 log cfu/g under in vivo conditions, whereas B. animalis KSp4 was not effective. But administration of bifidobacteria together with the prebiotic (as synbiotics) improved the bifidogenic effect by 1.4 log cfu/g of faeces. Supplementation of diet had almost no effect on the other determined groups of gut microflora. The obtained results showed the selective stimulation of faecal bifidobacteria by probiotics, prebiotic and synbiotics, a great bifidogenic effectiveness of oligofructose, confirmed proper selection of synbiotics and showed their higher effectiveness in relation to probiotics.

Introduction

The human gastrointestinal tract is a kinetic microecosystem that enables normal physiological functions of the host organism unless harmful and potentially pathogenic bacteria dominate it. Maintaining a proper equilibrium of the microflora may be ensured by systematic supplementation of the diet with probiotics, prebiotics or synbiotics. The most widely used definition of probiotic has been proposed by Fuller, 1989, Fuller, 1992, i.e. a live microbial feed supplement that beneficially affects the host animal by improving its intestinal microbial balance. Yet, for human nutrition Salminen et al. (1998) proposed the following definition: ‘a live microbial food ingredient that is beneficial to health’. Health advantages associated with probiotic intake are: alleviation of symptoms of lactose malabsorption, increase in natural resistance to infectious diseases of the intestinal tract, suppression of cancer, reduction in serum cholesterol concentrations, improved digestion, and stimulation of gastrointestinal immunity (Collins & Gibson, 1999), although, well-characterised strains with proven clinical effects are not numerous. Bacteria belonging to the genera Bifidobacterium and Lactobacillus are most often used as probiotic supplements for food. They exert only beneficial properties with regard to human health, such as inhibition of growth of exogeneous and/or harmful bacteria, stimulation of immune functions, anti-tumour properties, cholesterol reduction, aid in digestion and/or absorption of food ingredients/minerals and synthesis of vitamins (Gibson, 1998). The other commensal microflora additionally produces toxins, potential carcinogens, toxic H₂S and/or causes intestinal putrefaction, whereas activity of pathogenic bacteria results in diarrhoea or constipation, infections and harmful systemic effects.

In an analogy with probiotics, as living bacteria which added to food cross the upper gastrointestinal tract and colonise the gut, Delzenne and Roberfroid (1994) proposed the term prebiotics for the non-digestible oligosaccharides (NDO) used as food ingredients to modify the composition of endogenous gut microflora. The definition was developed by Gibson and Roberfroid (1995) who named a prebiotic ‘a nondigestible food ingredient that beneficially affects the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the colon, and thus improves host health’. Prebiotics comprise some oligosaccharides, especially fructo-oligosaccharides, which preferentially promote bifidobacteria growth in the large intestine. To date, the greatest scientific interest was focused on the nutritional and health benefits of oligofructose and inulin (Hartemink, 1997, Roberfroid and Milner, 1999). Consumption of appropriately selected probiotics as well as prebiotics may enhance the beneficial effect of each of them.

The synbiotics, as a combination of probiotics and prebiotics, have not been intensively studied to date. Some authors (Fooks et al., 1999, Roberfroid, 2000) suggest that they might improve the survival of the bacteria crossing the upper part of the gastrointestinal tract, thereby enhancing their effects in the large bowel. Perhaps their individual advantages might be additive or even synergistic but this statement needs to be qualified.

Generally, there is lack of studies on development of synbiotics from properly selected probiotics and prebiotics, followed by examination of their in vivo effectiveness. Unfortunately, they are mostly investigated separately. Therefore, the purpose of the present study was to examine different Bifidobacterium species preferences to defined fructo-oligosaccharides and fructans, to select stimulated strains and oligosaccharides enhancing their growth, develop their synbiotic pairs, as well as determine of effect of every factor on microecology of gut.